RESUMO
Asthma is a chronic inflammatory respiratory condition, characterized by variable airflow limitation, leading to clinical symptoms such as dyspnea and chest tightness. These symptoms result from an underlying inflammatory process. The ß2 agonists are bronchodilators prescribed for the relief of the disease. Nevertheless, their efficacy exhibits substantial interindividual variability. Currently, there is widespread recognition of the association between specific genetic variants, predominantly located within the ADRB2 and ADCY9 genes and their efficacy. This association, usually represented by the presence of non-synonymous single nucleotide polymorphisms (SNPs) have a strong impact in the protein functionality. The prevalence of these mutations varies based on the ethnic composition of the population and thus understanding the profiles of variability in different populations would contribute significantly to standardizing the use of these medications. In this study, we conducted a sequence-based genotyping of the relevant SNPs within the ADRB2 and ADCY9 genes in patients undergoing treatment with bronchodilators and/or corticosteroids at two healthcare facilities in the state of Rio de Janeiro, Brazil. We investigated the presence of c.46A>G, c.79C>G, c.252G>A, and c.491C>T SNPs within the ADRB2, and c.1320018 A>G within the ADCY9. Our results were in line with existing literature data with both for individuals in Brazil and Latin American.
RESUMO
Among non-tuberculous mycobacteria, Mycobacterium kansasii is one of the most pathogenic, able to cause pulmonary disease indistinguishable from tuberculosis in immunocompetent susceptible adults. The lack of animal models that reproduce human-like lung disease, associated with the necrotic lung pathology, impairs studies of M. kansasii virulence and pathogenicity. In this study, we examined the ability of the C57BL/6 mice, intratracheally infected with highly virulent M. kansasii strains, to produce a chronic infection and necrotic lung pathology. As a first approach, we evaluated ten M. kansasii strains isolated from Brazilian patients with pulmonary disease and the reference strain M. kansasii ATCC 12478 for virulence-associated features in macrophages infected in vitro; five of these strains differing in virulence were selected for in vivo analysis. Highly virulent isolates induced progressive lung disease in mice, forming large encapsulated caseous granulomas in later stages (120-150 days post-infection), while the low-virulent strain was cleared from the lungs by day 40. Two strains demonstrated increased virulence, causing premature death in the infected animals. These data demonstrate that C57BL/6 mice are an excellent candidate to investigate the virulence of M. kansasii isolates. We observed considerable heterogeneity in the virulence profile of these strains, in which the presence of highly virulent strains allowed us to establish a clinically relevant animal model. Comparing public genomic data between Brazilian isolates and isolates from other geographic regions worldwide demonstrated that at least some of the highly pathogenic strains isolated in Brazil display remarkable genomic similarities with the ATCC strain 12478 isolated in the United States 70 years ago (less than 100 SNPs of difference), as well as with some recent European clinical isolates. These data suggest that few pathogenic clones have been widely spread within M. kansasii population around the world.
RESUMO
Armadillos are specialist diggers and their burrows are used to find food, seek shelter and protect their pups. These burrows can also be shared with dozens of vertebrate and invertebrate species and; consequently, their parasites including the zoonotics. The aim of this study was to diagnose the presence of zoonotic parasites in four wild-caught armadillo species from two different Brazilian ecosystems, the Cerrado (Brazilian savanna) and the Pantanal (wetland). The investigated parasites and their correspondent diseases were: Toxoplasma gondii (toxoplasmosis), Trypanosoma cruzi (Chagas disease), Leishmania spp., (leishmaniasis), Paracoccidioides brasiliensis (Paracoccidioidomicosis) and Mycobacterium leprae (Hansen's disease). Forty-three free-living armadillos from Pantanal and seven road-killed armadillos from the Cerrado were sampled. Trypanosoma cruzi DTU TcIII were isolated from 2 out of 43 (4.65%) armadillos, including one of them also infected with Trypanosoma rangeli. Antibodies anti-T. gondii were detected in 13 out of 43 (30.2%) armadillos. All seven armadillos from Cerrado tested positive for P. brasiliensis DNA, in the lungs, spleen, liver fragments. Also, by molecular analysis, all 43 individuals were negative for M. leprae and Leishmania spp. Armadillos were infected by T. cruzi, T. rangeli, P. brasiliensis and presented seric antibodies to T. gondii, highlighting the importance of those armadillos could have in the epidemiology of zoonotic parasites.
Assuntos
Tatus , Doença de Chagas/veterinária , Leishmaniose/veterinária , Hanseníase/veterinária , Paracoccidioidomicose/veterinária , Toxoplasmose Animal/parasitologia , Zoonoses/microbiologia , Zoonoses/parasitologia , Animais , Brasil , Doença de Chagas/parasitologia , Feminino , Leishmania/isolamento & purificação , Leishmaniose/parasitologia , Hanseníase/microbiologia , Masculino , Mycobacterium leprae/isolamento & purificação , Paracoccidioides/isolamento & purificação , Paracoccidioidomicose/parasitologia , Especificidade da Espécie , Toxoplasma/isolamento & purificação , Trypanosoma cruzi/isolamento & purificaçãoRESUMO
ABSTRACT: Mycobacterium bovis is responsible for bovine and buffalo tuberculosis, an important zoonotic disease with global distribution. The knowledge of the distribution and the precise identification of this disease, including advanced diagnoses such as spoligotyping, allows choosing the best strategies to fight the diseases progress. The present work aimed to investigate mycobacterias presence, genotype their strains, and evaluate tuberculosis cases spatial distribution from suggestive lesions in carcasses of bovine and buffalo inspected in slaughterhouses under an official inspection regime in the state of Bahia, Brazil. The study investigated 453,417 animals. Among these, 31 (0.007%) from 17 municipalities were suspected of tuberculosis. Among the culture medium growth, 95% of these were categorized as alcohol-acid resistant bacilli (BAAR). All isolates were subjected to spoligotyping and 95% were confirmed as M. bovis (SB0120, SB0121, SB0852, SB0828, SB0295, SB0881, SB1648, SB6119, SB0140, SB1055). The strain SB0120 was the most prevalent, and this profile has been described in cases of human tuberculosis by M. bovis, highlighting the zoonotic potential of this profile. This study also identified strains never reported in Bahia, highlighting a distinctive pattern from other parts of Brazil, besides mixed infections. Besides, to identify strains never before described in the state, highlighting a distinctive pattern in Brazil (SB6119 and SB0852, respectively). An unpublished profile was identified and inserted in the international database (Mbovis.org), named SB2715.
RESUMO: O Mycobacterium bovis é o responsável pela tuberculose bovina e bubalina, doença zoonótica importante e com distribuição global. O conhecimento da distribuição e a identificação precisa dessa enfermidade, incluindo diagnósticos mais avançados como o spoligotyping, permite escolher as melhores estratégias de combate ao avanço da doença. O presente trabalho objetivou investigar a presença de micobactérias, genotipar suas estirpes e avaliar a distribuição espacial dos casos de tuberculose a partir de lesões sugestivas nas carcaças de bovinos e bubalinos inspecionadas em frigoríficos sob regime de inspeção oficial no estado da Bahia. Foram investigados 453.417 animais dentre os quais 31 (0,007%) foram suspeitos de doença e provenientes de 17 municípios. Após o crescimento em meio de cultura, 95% foram categorizados como bacilos álcool-ácido resistentes (BAAR). Todos os isolados foram submetidos à spoligotyping e 95% foram confirmados M. bovis (SB0120, SB0121, SB0852, SB0828, SB0295, SB0881, SB1648, SB6119, SB0140, SB1055). A cepa SB0120 foi a mais prevalente e este perfil vem sendo descrito na literatura com casos de tuberculose humana por M. bovis ressaltando o potencial zoonótico deste perfil. Este estudo também identificou cepas nunca relatadas no estado da Bahia, destacando um padrão distinto de outras partes do Brasil, além da existência de infecções mistas. Permitiu ainda relatar linhagens nunca antes descritas no estado com destaque para um padrão novo no Brasil (SB6119 e SB0852 respectivamente). Um perfil inédito identificado foi identificado e inserido no banco de dados internacional (Mbovis.org), nomeado SB2715.
RESUMO
Mycobacterium bovis is responsible for bovine and buffalo tuberculosis, an important zoonotic disease with global distribution. The knowledge of the distribution and the precise identification of this disease, including advanced diagnoses such as spoligotyping, allows choosing the best strategies to fight the disease's progress. The present work aimed to investigate mycobacteria's presence, genotype their strains, and evaluate tuberculosis cases' spatial distribution from suggestive lesions in carcasses of bovine and buffalo inspected in slaughterhouses under an official inspection regime in the state of Bahia, Brazil. The study investigated 453,417 animals. Among these, 31 (0.007%) from 17 municipalities were suspected of tuberculosis. Among the culture medium growth, 95% of these were categorized as alcohol-acid resistant bacilli (BAAR). All isolates were subjected to spoligotyping and 95% were confirmed as M. bovis (SB0120, SB0121, SB0852, SB0828, SB0295, SB0881, SB1648, SB6119, SB0140, SB1055). The strain SB0120 was the most prevalent, and this profile has been described in cases of human tuberculosis by M. bovis, highlighting the zoonotic potential of this profile. This study also identified strains never reported in Bahia, highlighting a distinctive pattern from other parts of Brazil, besides mixed infections. Besides, to identify strains never before described in the state, highlighting a distinctive pattern in Brazil (SB6119 and SB0852, respectively). An unpublished profile was identified and inserted in the international database (Mbovis.org), named SB2715.(AU)
O Mycobacterium bovis é o responsável pela tuberculose bovina e bubalina, doença zoonótica importante e com distribuição global. O conhecimento da distribuição e a identificação precisa dessa enfermidade, incluindo diagnósticos mais avançados como o spoligotyping, permite escolher as melhores estratégias de combate ao avanço da doença. O presente trabalho objetivou investigar a presença de micobactérias, genotipar suas estirpes e avaliar a distribuição espacial dos casos de tuberculose a partir de lesões sugestivas nas carcaças de bovinos e bubalinos inspecionadas em frigoríficos sob regime de inspeção oficial no estado da Bahia. Foram investigados 453.417 animais dentre os quais 31 (0,007%) foram suspeitos de doença e provenientes de 17 municípios. Após o crescimento em meio de cultura, 95% foram categorizados como bacilos álcool-ácido resistentes (BAAR). Todos os isolados foram submetidos à spoligotyping e 95% foram confirmados M. bovis (SB0120, SB0121, SB0852, SB0828, SB0295, SB0881, SB1648, SB6119, SB0140, SB1055). A cepa SB0120 foi a mais prevalente e este perfil vem sendo descrito na literatura com casos de tuberculose humana por M. bovis ressaltando o potencial zoonótico deste perfil. Este estudo também identificou cepas nunca relatadas no estado da Bahia, destacando um padrão distinto de outras partes do Brasil, além da existência de infecções mistas. Permitiu ainda relatar linhagens nunca antes descritas no estado com destaque para um padrão novo no Brasil (SB6119 e SB0852 respectivamente). Um perfil inédito identificado foi identificado e inserido no banco de dados internacional (Mbovis.org), nomeado SB2715.(AU)
Assuntos
Animais , Bovinos , Búfalos/genética , Mycobacterium bovis , Bovinos/genética , Zoonoses , Técnicas de GenotipagemRESUMO
Molecular epidemiology investigations are notoriously challenging in the leprosy field mainly because the inherent characteristics of the disease as well as its yet uncultivated causative agents, Mycobacterium leprae and M. lepromatosis. Despite significant developments in understanding the biology of leprosy bacilli through genomic approaches, the exact mechanisms of transmission is still unclear and the factors underlying pathological variation of the disease in different patients remain as major gaps in our knowledge about leprosy. Despite these difficulties, the last two decades have seen the development of genotyping procedures based on PCR-sequencing of target loci as well as by the genome-wide analysis of an increasing number of geographically diverse isolates of leprosy bacilli. This has provided a foundation for molecular epidemiology studies that are bringing a better understanding of strain evolution associated with ancient human migrations, and phylogeographical insights about the spread of disease globally. This review discusses the advantages and drawbacks of the main tools available for molecular epidemiological investigations of leprosy and summarizes various methods ranging from PCR-based genotyping to genome-typing techniques. We also describe their main applications in analyzing the short-range and long-range transmission of the disease. Finally, we summarise the current gaps and challenges that remain in the field of molecular epidemiology of leprosy.
Assuntos
Hanseníase/epidemiologia , Hanseníase/microbiologia , Mycobacterium leprae/genética , Genes Bacterianos , Genoma Bacteriano , Genômica/métodos , Humanos , Hanseníase/tratamento farmacológico , Hanseníase/transmissão , Epidemiologia Molecular , Mycobacterium leprae/efeitos dos fármacos , Filogenia , Vigilância em Saúde PúblicaRESUMO
BACKGROUND: Leprosy has been treated with multidrug therapy, which has been distributed for free across the globe and regarded as highly efficient. However, the impossibility of growing Mycobacterium leprae in axenic media has historically impaired assessments of M. leprae resistance, a parameter only recently detectable through molecular methods. METHODS: A systematic, population-based search for M. leprae resistance in suspected leprosy relapse cases and contacts was performed in Prata Village, an isolated, hyperendemic, former leprosy colony located in the Brazilian Amazon. Results led to an extended active search involving the entire Prata population. Confirmed leprosy cases were investigated for bacterial resistance using a combination of in vivo testing and direct sequencing of resistance genes folP1, rpoB, and gyrA. A molecular epidemiology analysis was performed using data from 17 variable number tandem repeats (VNTR). RESULTS: Mycobacterium leprae was obtained from biopsies of 37 leprosy cases (18 relapses and 19 new cases): 16 (43.24%) displayed drug-resistance variants. Multidrug resistance to rifampicin and dapsone was observed in 8 relapses and 4 new cases. Single resistance to rifampicin was detected in 1 new case. Resistance to dapsone was present in 2 relapses and 1 new case. Combined molecular resistance and VNTR data revealed evidence of intra-familial primary transmission of resistant M. leprae. CONCLUSIONS: A comprehensive, population-based systematic approach to investigate M. leprae resistance in a unique population revealed an alarming scenario of the emergence and transmission of resistant strains. These findings may be used for the development of new strategies for surveillance of drug resistance in other populations.
Assuntos
Hanseníase , Preparações Farmacêuticas , Brasil/epidemiologia , Farmacorresistência Bacteriana , Quimioterapia Combinada , Humanos , Hansenostáticos/farmacologia , Hansenostáticos/uso terapêutico , Hanseníase/tratamento farmacológico , Hanseníase/epidemiologia , Testes de Sensibilidade Microbiana , Mycobacterium leprae/genéticaRESUMO
Mycobacterium kansasii is an opportunistic pathogen and one of the most commonly encountered species in individuals with lung disease. We here report the complete genome sequence of 12 clinical isolates of M. kansasii from patients with pulmonary disease in Brazil.
Assuntos
Genoma Bacteriano , Genótipo , Pneumopatias/microbiologia , Infecções por Mycobacterium não Tuberculosas/microbiologia , Mycobacterium kansasii/genética , Brasil , Gráficos por Computador , DNA Bacteriano , Genômica , Humanos , Mycobacterium kansasii/isolamento & purificação , Polimorfismo de Fragmento de RestriçãoRESUMO
Tuberculosis (TB) remains a major public health problem in the world and Brazil is among the countries with the highest incidence and prevalence rates, and Rio Grande do Sul, a Brazilian state, occupy a prominent position. Multidrug-resistant Mycobacterium tuberculosis (MDR-TB) further aggravates this scenario, making it more difficult to treat and control the disease. Isoniazid monoresistance (IMR) may increase the risk of progression to MDR-TB and treatment failure. However, most drug resistance molecular tests only focus on detecting rifampicin (RIF) resistance.In the present study, we characterized a total of 63 drug resistant isolates of M. tuberculosis (35 MDR, 26 IMR and two isolates monoresistant to rifampicin [RMR]) of the Rio Grande do Sul state by MIRU-VNTR (24 loci), spoligotyping, presence of RDRio, fbpC103, pks15/1 and sequencing of the katG, rpoB and inhA genes. We observed a higher proportion of the LAM family 30/63 (47.61%). In IMR, mutations were found in the katG gene (98% at codon 315) in 72.5%, and mutations in the promoter region of the inhA gene in 6.25% of the isolates. In MDR-TB and RMR-TB isolates, 92.1% had mutations in the rpoB gene (57% at codon 531). The presence of a 12 bp insertion between codons 516 and 517 of the rpoB gene in MDR-TB isolates was found in five isolates. In conclusion, we observed that the highest frequency of IMR-TB and MDR-TB strains belong to the LAM and Haarlem genotypes in Rio Grande do Sul state. A significant number of isolates previously characterized as Mycobacterium pinnipedi2 through spoligotyping were found to belong to the M. tuberculosis LAM family. This was responsible for a number of significant cases and the molecular profile of this strain and the pattern of mutations related to drug resistance were analyzed. These findings may contribute to a better understanding about the spread of M. tuberculosis resistant in southern of Brazil.
Assuntos
Antituberculosos/farmacologia , Isoniazida/farmacologia , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/genética , Tuberculose/microbiologia , Técnicas de Tipagem Bacteriana/métodos , Farmacorresistência Bacteriana/genética , Farmacorresistência Bacteriana Múltipla/genética , Genes Bacterianos/genética , Variação Genética , Genótipo , Humanos , Testes de Sensibilidade Microbiana/métodos , Repetições Minissatélites/genética , Mycobacterium tuberculosis/classificação , Fenótipo , Tuberculose/tratamento farmacológico , Tuberculose Resistente a Múltiplos Medicamentos/tratamento farmacológico , Tuberculose Resistente a Múltiplos Medicamentos/microbiologiaRESUMO
Leprosy is a chronic human disease caused by the yet-uncultured pathogen Mycobacterium leprae. Although readily curable with multidrug therapy (MDT), over 200,000 new cases are still reported annually. Here, we obtain M. leprae genome sequences from DNA extracted directly from patients' skin biopsies using a customized protocol. Comparative and phylogenetic analysis of 154 genomes from 25 countries provides insight into evolution and antimicrobial resistance, uncovering lineages and phylogeographic trends, with the most ancestral strains linked to the Far East. In addition to known MDT-resistance mutations, we detect other mutations associated with antibiotic resistance, and retrace a potential stepwise emergence of extensive drug resistance in the pre-MDT era. Some of the previously undescribed mutations occur in genes that are apparently subject to positive selection, and two of these (ribD, fadD9) are restricted to drug-resistant strains. Finally, nonsense mutations in the nth excision repair gene are associated with greater sequence diversity and drug resistance.
Assuntos
Anti-Infecciosos/farmacologia , Farmacorresistência Bacteriana/genética , Mycobacterium leprae/efeitos dos fármacos , Filogenia , Códon sem Sentido , DNA Bacteriano/química , Genoma Bacteriano , Humanos , Testes de Sensibilidade Microbiana , Mycobacterium leprae/genética , Mycobacterium leprae/isolamento & purificaçãoAssuntos
Técnicas de Diagnóstico Molecular/métodos , Mycobacterium tuberculosis/isolamento & purificação , Reação em Cadeia da Polimerase , Tuberculose/diagnóstico , Proteínas de Bactérias/genética , DNA Bacteriano/genética , Variação Genética , Genótipo , Mycobacterium tuberculosis/genética , Federação Russa , Fator sigma/genética , Fatores de Tempo , Tuberculose/microbiologia , Tuberculose Resistente a Múltiplos Medicamentos/diagnóstico , Tuberculose Resistente a Múltiplos Medicamentos/microbiologiaRESUMO
Mycobacterium kansasii is an opportunistic pathogen and one of the most commonly encountered species in individuals with lung disease. We here report the complete genome sequence of 12 clinical isolates of M. kansasii from patients with pulmonary disease in Brazil.
Assuntos
DNA Bacteriano , Genoma Bacteriano/genética , Mycobacterium kansasii/genética , Gráficos por ComputadorRESUMO
Leprosy is a chronic human disease caused by the yet-uncultured pathogen Mycobacterium leprae. Although readily curable with multidrug therapy (MDT), over 200,000 new cases are still reported annually. Here, we obtain M. leprae genome sequences from DNA extracted directly from patients' skin biopsies using a customized protocol. Comparative and phylogenetic analysis of 154 genomes from 25 countries provides insight into evolution and antimicrobial resistance, uncovering lineages and phylogeographic trends, with the most ancestral strains linked to the Far East. In addition to known MDT-resistance mutations, we detect other mutations associated with antibiotic resistance, and retrace a potential stepwise emergence of extensive drug resistance in the pre-MDT era. Some of the previously undescribed mutations occur in genes that are apparently subject to positive selection, and two of these (ribD, fadD9) are restricted to drug-resistant strains. Finally, nonsense mutations in the nth excision repair gene are associated with greater sequence diversity and drug resistance.
Assuntos
Humanos , Filogenia , DNA Bacteriano/química , Testes de Sensibilidade Microbiana , Genoma Bacteriano , Códon sem Sentido , Farmacorresistência Bacteriana/genética , Anti-Infecciosos/farmacologia , Mycobacterium leprae/isolamento & purificação , Mycobacterium leprae/efeitos dos fármacos , Mycobacterium leprae/genéticaRESUMO
Leprosy is endemic in large part of Brazil with 28,761 new patients in 2015, the second largest number worldwide and reaches 9/10.000 in highly endemic regions and 2.7/10.000 in the city of Fortaleza, Ceará, Northeast Brazil. For better understanding of risk factors for leprosy transmission, we conducted an epidemiologic study supplemented by 17 locus VNTR and SNP 1-4 typing of Mycobacterium leprae in skin biopsy samples from new multibacillary (MB) patients diagnosed at a reference center in 2009 and 2010. Among the 1,519 new patients detected during the study period, 998 (65.7%) were MB and we performed DNA extraction and genotyping on 160 skin biopsy samples, resulting in 159 (16%) good multilocus VNTR types. Thirty-eight of these patients also provided VNTR types from M. leprae in nasal swabs. The SNP-Type was obtained for 157 patients and 87% were of type 4. Upon consideration all VNTR markers, 156 different genotypes and three pairs with identical genotypes were observed; no epidemiologic relation could be observed between individuals in these pairs. Considerable variability in differentiating index (DI) was observed between the different markers and the four with highest DI [(AT)15, (TA)18, (AT)17 and (GAA)21] frequently demonstrated differences in copy number when comparing genotypes from both type of samples. Excluding these markers from analysis resulted in 83 genotypes, 20 of which included 96 of the patients (60.3%). These clusters were composed of two (n = 8), three (n = 6), four (n = 1), five (n = 2), six (n = 1), 19 (n = 1) and 23 (n = 23) individuals and suggests that recent transmission is contributing to the maintenance of leprosy in Fortaleza. When comparing epidemiological and clinical variables among patients within clustered or with unique M. leprae genotypes, a positive bacterial index in skin biopsies and knowledge of working with someone with the disease were significantly associated with clustering. A tendency to belong to a cluster was observed with later notification of disease (mean value of 3.4 months) and having disability grade 2. A tendency for lack of clustering was observed for patients who reported to have lived with another leprosy case but this might be due to lack of inclusion of household contacts in the study. Although clusters were spread over the city, kernel analysis revealed that some of the patients belonging to the two major clusters were spatially related to some neighborhoods that report poverty and high disease incidence in children. Finally, inclusion of genotypes from nasal swabs might be warranted. A major limitation of the study is that sample size of 160 patients from a two year period represents only 15% of the new patients and this could have weakened statistical outcomes. This is the first molecular epidemiology study of leprosy in Brazil and although the high clustering level suggests that recent transmission is the major cause of disease in Fortaleza; the existence of two large clusters needs further investigation.
Assuntos
Hanseníase/transmissão , Mycobacterium leprae/genética , Brasil/epidemiologia , Análise por Conglomerados , Genótipo , Geografia , Humanos , Hanseníase/microbiologia , Repetições Minissatélites/genética , Epidemiologia Molecular , Fatores de Risco , Análise EspacialRESUMO
Salmonella enterica subsp. enterica serovar Typhimurium is a surveyed worldwide serotype with well-characterized genomes for several different strains. In Brazil, very few studies have submitted whole-genome sequences to GenBank. This genome may be useful to analyze the genetic mechanisms comparable to those of other related studies conducted in Brazil and globally.
RESUMO
Background: This study compares the strains of genotypes of M. leprae from nasal secretions (NS) and skin biopsy (SB) in the same patient, supplementing conventional epidemiology to gain insight into the infection of leprosy in Fortaleza, Brazil. Methods: The sample consisted of 38 newly diagnosed leprosy patients attending the National Reference Center of Dermatology Dona Libania (CDERM), in Fortaleza, who tested positive for M. leprae by PCR in DNA extracts of nasal secretions. DNA was also extracted from skin biopsy (SB) scrapings of each patient and used for multiplex PCR amplification of M. leprae VNTR loci. The number of repeats at 15 loci were determined by the fragment length analysis method. Results: Locus VNTR genotypes were achieved in 38 NS, and in 38 SB specimens. M. leprae strains differed in their genotypes in paired specimens in all but two of 38 patients. The genotype similarity in the remainder ranged from 53% to 87%. Conclusion: M. leprae 15 VNTR loci genotypes of paired nasal and biopsy skin samples from five patients were identical, while as many as seven loci differed in the 33 other patients. When the NS and biopsy genotypes were pooled and compared, it was found that there was a great variability among different VNTR markers. It is important to investigate other molecular markers suitable for typing genetic variations of the bacilli.
Assuntos
Biópsia/métodos , Hanseníase/microbiologia , Repetições Minissatélites , Mycobacterium leprae/genética , Nariz/microbiologia , Pele/patologia , Brasil/epidemiologia , Estudos Transversais , DNA Bacteriano/genética , Doenças Endêmicas , Variação Genética , Genótipo , Humanos , Hanseníase/diagnóstico , Mycobacterium leprae/classificação , Mycobacterium leprae/isolamento & purificação , Reação em Cadeia da Polimerase , Polimorfismo de Nucleotídeo Único , Pele/microbiologiaRESUMO
We conducted a population-based study of tuberculosis (TB) cases in Dourados, Brazil, to assess the relationship between incarceration and TB in the general population. Incarceration was associated with TB in an urban population; 54% of Mycobacterium tuberculosis strains were related to strains from persons in prisons. TB control in prisons is critical for reducing disease prevalence.
Assuntos
Reservatórios de Doenças , Mycobacterium tuberculosis/genética , Prisões , Tuberculose/epidemiologia , Tuberculose/transmissão , Brasil/epidemiologia , Estudos de Casos e Controles , Feminino , Humanos , Masculino , Tipagem Molecular , Mycobacterium tuberculosis/classificação , Vigilância da População , Prevalência , Fatores de RiscoRESUMO
Post-mortem bacterial culture and specific biochemical tests are currently performed to characterize the etiologic agent of bovine tuberculosis. Cultures take up to 90 days to develop. A diagnosis by molecular tests such as PCR can provide fast and reliable results while significantly decreasing the time of confirmation. In the present study, a nested-PCR system, targeting rv2807, with conventional PCR followed by real-time PCR, was developed to detect Mycobacterium tuberculosis complex (MTC) organisms directly from bovine and bubaline tissue homogenates. The sensitivity and specificity of the reactions were assessed with DNA samples extracted from tuberculous and non-tuberculous mycobacteria, as well as other Actinomycetales species and DNA samples extracted directly from bovine and bubaline tissue homogenates. Regarding the analytical sensitivity, DNA of the M. bovis AN5 strain was detected up to 1.5 pg by nested-PCR, whereas DNA of M. tuberculosis H37Rv strain was detected up to 6.1 pg. The nested-PCR system showed 100% analytical specificity for MTC when tested with DNA of reference strains of non-tuberculous mycobacteria and closely-related Actinomycetales. A clinical sensitivity level of 76.7% was detected with tissues samples positive for MTC by means of the culture and conventional PCR. A clinical specificity of 100% was detected with DNA from tissue samples of cattle with negative results in the comparative intradermal tuberculin test. These cattle exhibited no visible lesions and were negative in the culture for MTC. The use of the nested-PCR assay to detect M. tuberculosis complex in tissue homogenates provided a rapid diagnosis of bovine and bubaline tuberculosis.
Assuntos
Técnicas de Diagnóstico Molecular/métodos , Mycobacterium bovis/isolamento & purificação , Mycobacterium tuberculosis/isolamento & purificação , Patologia Molecular/métodos , Reação em Cadeia da Polimerase/métodos , Tuberculose Bovina/diagnóstico , Medicina Veterinária/métodos , Animais , Búfalos , Bovinos , Mycobacterium bovis/genética , Mycobacterium tuberculosis/genética , Sensibilidade e Especificidade , Fatores de Tempo , Tuberculose Bovina/microbiologiaRESUMO
BACKGROUND: Multidrug resistance is a critical factor in tuberculosis control. To gain better understanding of multidrug resistant tuberculosis in Brazil, a retrospective study was performed to compare genotypic diversity and drug resistance associated mutations in Mycobacterium tuberculosis isolates from a national reference center. METHODS AND FINDINGS: Ninety-nine multidrug resistant isolates from 12 Brazilian states were studied. Drug-resistance patterns were determined and the rpoB and katG genes were screened for mutations. Genotypic diversity was investigated by IS6110-RFLP and Luminex 47 spoligotyping. Mutations in rpoB and katG were seen in 91% and 93% of the isolates, respectively. Codon 315 katG mutations occurred in 82.8% of the isolates with a predominance of the Ser315Thr substitution. Twenty-five isolates were clustered in 11 groups with identical IS6110-RFLP patterns while 74 showed unique patterns with no association between mutation frequencies or susceptibility profiles. The most prevalent spoligotyping lineages were LAM (47%), T (17%) and Haarlen (12%). The Haarlen lineage showed a higher frequency of codon 516 rpoB mutations while codon 531 mutations prevailed in the other isolates. CONCLUSIONS: Our data suggest that there were no major multidrug resistant M. tuberculosis strains transmitted among patients referred to the reference center, indicating an independent acquisition of resistance. In addition, drug resistance associated mutation profiles were well established among the main spoligotyping lineages found in these Brazilian multidrug resistant isolates, providing useful data for patient management and treatment.
Assuntos
Proteínas de Bactérias/genética , Catalase/genética , Farmacorresistência Bacteriana Múltipla/genética , Mutação/genética , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/isolamento & purificação , Tuberculose Resistente a Múltiplos Medicamentos/genética , Adulto , Técnicas de Tipagem Bacteriana , Brasil/epidemiologia , Análise por Conglomerados , RNA Polimerases Dirigidas por DNA , Demografia , Feminino , Humanos , Incidência , Masculino , Mycobacterium tuberculosis/classificação , Filogenia , Polimorfismo de Fragmento de Restrição , Estudos Retrospectivos , Tuberculose Resistente a Múltiplos Medicamentos/epidemiologiaRESUMO
Post-mortem bacterial culture and specific biochemical tests are currently performed to characterize the etiologic agent of bovine tuberculosis. Cultures take up to 90 days to develop. A diagnosis by molecular tests such as PCR can provide fast and reliable results while significantly decreasing the time of confirmation. In the present study, a nested-PCR system, targeting rv2807, with conventional PCR followed by real-time PCR, was developed to detect Mycobacterium tuberculosis complex (MTC) organisms directly from bovine and bubaline tissue homogenates. The sensitivity and specificity of the reactions were assessed with DNA samples extracted from tuberculous and non-tuberculous mycobacteria, as well as other Actinomycetales species and DNA samples extracted directly from bovine and bubaline tissue homogenates. Regarding the analytical sensitivity, DNA of the M. bovis AN5 strain was detected up to 1.5 pg by nested-PCR, whereas DNA of M. tuberculosis H37Rv strain was detected up to 6.1 pg. The nested-PCR system showed 100% analytical specificity for MTC when tested with DNA of reference strains of non-tuberculous mycobacteria and closely-related Actinomycetales. A clinical sensitivity level of 76.7% was detected with tissues samples positive for MTC by means of the culture and conventional PCR. A clinical specificity of 100% was detected with DNA from tissue samples of cattle with negative results in the comparative intradermal tuberculin test. These cattle exhibited no visible lesions and were negative in the culture for MTC. The use of the nested-PCR assay to detect M. tuberculosis complex in tissue homogenates provided a rapid diagnosis of bovine and bubaline tuberculosis.